TY - JOUR
T1 - Detection of histidine kinases via a filter-based assay and reverse-phase thin-layer chromatographic phosphoamino acid analysis
AU - Tan, E.
AU - Zu, X.L.
AU - Yeoh, George
AU - Besant, Paul
AU - Attwood, Paul
PY - 2003
Y1 - 2003
N2 - The methods that detect histidine phosphorylation have largely been either laborious or difficult to apply quantitatively. The major difficulty in assessing for its presence is its alkali-stable, acid-labile nature. While an assay that detects alkali-stable phosphorylation has been developed, it does not distinguish phosphohistidine from other alkali-stable phosphoamino acids. Using this established method, we extend the assay to facilitate the specific detection of phosphohistidine. We use the acid-lability of phosphohistidine as a defining feature in our approach for its detection. In addition, reverse-phase thin-layer chromatography was utilized to conclusively demonstrate the viability of the conditions that we implement in the assay for the selective detection of phosphohistidine. In summary, this report describes a rapid filter-based kinase assay that quantitatively measures histidine kinase activity, even in the presence of tyrosine kinase activity. (C) 2003 Elsevier Inc. All rights reserved.
AB - The methods that detect histidine phosphorylation have largely been either laborious or difficult to apply quantitatively. The major difficulty in assessing for its presence is its alkali-stable, acid-labile nature. While an assay that detects alkali-stable phosphorylation has been developed, it does not distinguish phosphohistidine from other alkali-stable phosphoamino acids. Using this established method, we extend the assay to facilitate the specific detection of phosphohistidine. We use the acid-lability of phosphohistidine as a defining feature in our approach for its detection. In addition, reverse-phase thin-layer chromatography was utilized to conclusively demonstrate the viability of the conditions that we implement in the assay for the selective detection of phosphohistidine. In summary, this report describes a rapid filter-based kinase assay that quantitatively measures histidine kinase activity, even in the presence of tyrosine kinase activity. (C) 2003 Elsevier Inc. All rights reserved.
U2 - 10.1016/j.ab.2003.08.035
DO - 10.1016/j.ab.2003.08.035
M3 - Article
SN - 0003-2697
VL - 323
SP - 122
EP - 126
JO - Analytical Biochemistry
JF - Analytical Biochemistry
ER -